This work examines the inter-relationship between the unbound drug fractions in blood and brain homogenate, passive membrane permeability, P-glycoprotein (Pgp) efflux ratio, and log octanol/water partition coefficients (cLogP) in determining the extent of central nervous system (CNS) penetration observed in vivo. The present results demonstrate that compounds often considered to be Pgp substrates in rodents (efflux ratio greater than 5 in multidrug resistant Madin-Darby canine kidney cells) with poor passive permeability may still exhibit reasonable CNS penetration in vivo; i.e., where the unbound fractions and nonspecific tissue binding act as a compensating force. In these instances, the efflux ratio and in vitro blood-brain partition ratio may be used to predict the in vivo blood-brain ratio. This relationship may be extended to account for the differences in CNS penetration observed in vivo between mdr1a/b wild type and knockout mice. In some instances, cross-species differences that might initially seem to be related to differing transporter expression can be rationalized from knowledge of unbound fractions alone. The results presented in this article suggest that the information exists to provide a coherent picture of the nature of CNS penetration in the drug discovery setting, allowing the focus to be shifted away from understanding CNS penetration toward the more important aspect of understanding CNS efficacy.Within the modern drug discovery paradigm, drug metabolism and pharmacokinetics (DMPK) play an integral role in the process of compound selection and progression. Much of the impact of DMPK has been caused by its transformation from a largely descriptive discipline to that of a predictive science, fuelled by advances in bioanalysis and in vitro techniques. Hence discovery DMPK provides a powerful means for assessing the risks of taking potential assets into development.Nevertheless, the development of molecules targeted at the central nervous system (CNS) remains a significant challenge caused by the increased regulation and protection afforded to the brain over other organs of the body. The major knowledge gaps are 1) understanding the physicochemical features that determine CNS penetration, 2) understanding the impact of the blood-brain barrier (BBB) on CNS uptake, and 3) providing a coherent measure of CNS penetration that can be related to drug efficacy. Regarding the latter point, although it is important to develop a link between the pharmacokinetics of a molecule and the biophase, arguably the critical issue is one of sufficient access of free drug to the requisite site of action.Numerous models and measures of CNS uptake are available to assist in the search for centrally active agents. In situ brain perfusion techniques have highlighted the good correlation between increasing lipophilicity and CNS permeability. Polar drugs that are subject to paracellular absorption such as atenolol (logD oct,7.4 Ϫ2.1; Artursson, 1990) and sumatriptan (logD oct,7.4 Ϫ1.5;Pascual and Munoz, 2005) show...

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1 Understanding the mechanisms underlying the analgesic effect of new cyclooxygenase inhibitors is essential to identify dosing requirements in early stages of drug development. Accurate extrapolation to humans of in vitro and in vivo findings in preclinical species is needed to optimise dosing regimen in inflammatory conditions. 2 The current investigation characterises the inhibition of prostaglandin E2 (PGE 2 ) and thromboxane B2 (TXB 2 ) by naproxen in vitro and in vivo in rat and human blood. The inhibition of PGE 2 in the absence or presence of increasing concentrations of naproxen (10 À8 -10 À1 M) was measured by ex vivo whole blood stimulation with LPS, whereas inhibition of TXB 2 was measured in serum following blood clotting. In further experiments, inhibition of PGE 2 and TXB 2 levels was also assessed ex vivo in animals treated with naproxen (2.5, 10, 25 mg kg À1 ). Subsequently, pharmacokinetic (PK)/pharmacodynamics (PD) modelling of in vitro and in vivo data was performed using nonlinear mixed effects in NONMEM (V). 3 Inhibition of PGE 2 and TXB 2 was characterised by a sigmoid E max model. The exposure-response relationships in vitro and in vivo were of the same order of magnitude in both species. IC 80 estimates obtained in vitro were similar for PGE 2 inhibition (130.8711 and 131.9719 10 À6 M, mean7s.d. for humans and rats, respectively), but slightly different for TXB 2 inhibition (103.9715 and 151.4740 10 À6 M, mean7s.d. for humans and rats, respectively, Po 0.05). These differences, however, may not be biologically relevant. 4 The results confirm the value of exposure-effect relationships determined in vitro as a means to predict the pharmacological activity in vivo. This analysis also highlights the need to parameterise concentration-effect relationships in early drug development, as indicated by the estimates of IC 80 for PGE 2 and TXB 2 inhibition.

We studied the oxidation capacity in liver biopsies of a series of extensive metabolizers (n = 10) and poor metabolizers (n = 2) as identified by in vivo phenotyping with dextromethorphan. Codeine and dextromethorphan were used as probe drugs in vitro. The data were compared with the contents of cytochrome P-450IID1 as quantitated by Western immunoblotting by use of a specific monoclonal antibody (MAb 114/2). The O-demethylation of codeine was highly correlated with the O-demethylation of dextromethorphan (r = 0.90). The N-demethylation of codeine was catalyzed at a considerably higher rate than the O-demethylation. The N-demethylation to O-demethylation ratio of codeine was 46 in the poor metabolizer and, on average, 6.2 (range, 2.6 to 11) in the extensive metabolizers, respectively. The band intensity in Western blots correlated with the rate of O-demethylation of codeine (r = 0.95) and of dextromethorphan (r = 0.88) in the extensive metabolizers. The comeasurement of the O-demethylation and N-demethylation of codeine may provide a tool with which to phenotype individuals in vitro with respect to the polymorphism of the cytochrome P-450IID1.

Changes in paw pressure threshold seem to best reflect the anti-hyperalgesic properties of COX-inhibitors with enough sensitivity to enable estimation of the dose-exposure-response curve.